In silico identification and experimental validation of cellular uptake by a new cell penetrating peptide P1 derived from MARCKS

Abstract

Viral vectors for vaccine delivery are challenged by recently reported safety issues like immunogenicity and risk for cancer development, and thus there is a growing need for the development of non-viral vectors. Cell penetrating peptides (CPPs) are non-viral vectors that can enter plasma membranes efficiently and deliver a broad range of cargoes. Our bioinformatic prediction and wet-lab validation data suggested that peptide P1 derived from MARCKS protein phosphorylation site domain is a new potential CPP candidate. We found that peptide P1 can efficiently internalize into various cell lines in a concentration-dependent manner. Receptor-mediated endocytosis pathway is the major mechanism of P1 penetration, although P1 also directly penetrates the plasma membrane. We also found that peptide P1 has low cytotoxicity in cultured cell lines as well as mouse red blood cells. Furthermore, peptide P1 not only can enter into cultured cells itself, but it also can interact with plasmid DNA and mediate the functional delivery of plasmid DNA into cultured cells, even in hard-to-transfect cells. Combined, these findings indicate that P1 may be a promising vector for efficient intracellular delivery of bioactive cargos.

Cell-Permeable PROTAC Degraders against KEAP1 Efficiently Suppress Hepatic Stellate Cell Activation through the Antioxidant and Anti-Inflammatory Pathway

“…FITC-labeled KKP1 in the cell has the strongest fluorescence intensity during the initial 1 and 2 h of treatment and gradually decreased after 4, 8, 16, and 24 h (Figures D,E and S3). Our previous publications suggest that a penetration enhancer − can significantly enhance the penetration efficiency of the well-known different CPPs including hPP3, hPP10, , MT23, Scp01-b, Dot1l, P2, and P1 . We wanted to address whether the penetration efficiency of KKP can further be enhanced.…”

“…Thus, effective therapeutic strategies are still in urgent demand. We successfully developed a variety of delivery vectors of CPPs to deliver biotherapeutics into intracellular in vitro cultured cells and the in vivo mouse model. − ,,,− ,,, Thus, with our combined expertise, we attempted to develop a CPP-based PROTAC.…”

“…Our previous publications suggest that a penetration enhancer 43−45 can significantly enhance the penetration efficiency of the wellknown different CPPs including hPP3, 23 hPP10, 21,46 MT23, 47 Scp01-b, 48 Dot1l, 49 P2, 50 and P1. 51 We wanted to address whether the penetration efficiency of KKP can further be enhanced. Therefore, we used treatments with or without 5% DMSO in the HSC-T6 cells and then examined the efficiency of KKP (Figure 3F,G).…”

Improved transfer efficiency of supercharged 36 + GFP protein mediate nucleic acid delivery

“…Our studies also suggested although 36 + GFP can significantly enter cells and tissues compared with GFP protein, the penetration efficiency of 36 + GFP is still limited. Following our previous studies (Ma et al., 2015 ; Wang et al., 2016a , 2016b , 2016c ; Zhou et al., 2017 ; Ding et al., 2019 ; Zhang et al., 2019 ; Chen et al., 2021 ; Guo et al., 2021 ), we conducted 5% DMSO treatment in cultured cells, and found that the efficiency of 36 + GFP mediated transduction itself and its mediated transfection were significantly increase thus these results are consistent with our previous published studies. These data indicated that penetration enhancer application is another alternative approach to enhance the penetration and relative delivery efficiency of peptide or supercharged proteins- based delivery vectors.…”

Applications of Metallic Nanoparticles in the Skin Cancer Treatment

“…Nanoparticles. NPs are composed of particles dispersed below nanometers or solid particles with a size span of 10-1000 nm with biomimetic attributes [31,32]. Biomimetic attributes increase in combination with the high surfaceto-volume proportion.…”